Sialic acids are responsible for important physiological actions such as intercellular transmissions, cytoplasmic interactions, and cellular adhesions. Existences of wide variety of different cell-surface sialic acids are known, and they are regulated in processes of generations, differenciations, and transformations of oncogenes. Sialic acids are ubiquitous in the oligosaccharide side chains of glycoconjugates of a wide variety of animals (Varki, A., Curr. Opin. Cell. Biol. 4, pp.257-266, 1992).
Sialic acids exist at the end of hydrocarbon groups of glycoproteins and glycolipids. Sialic acids are enzymatically introduced to these positions from CMP-Sia during post-translation processes. For example, three sequential types, i.e., Sia.alpha. 2,6Gal, Sia.alpha. 2,3Gal, and Sia.alpha. 2,6GalNAc, commonly exist in glycoproteins (Hakomori, S., Ann. Rev. Biochem., 50, 733-764, 1981), and two sequential types, i.e. Sia.alpha. 2,3Gal and Sia.alpha. 2,8Sia, are frequently observed in ganglyosides (Fishman, P., and Brady, R. O., Science, 194, 906-915, 1976).
Enzymes responsible for the above-mentioned enzymatic introductions of sialic acids (i.e. sialyltransfers) are glycosyltransferases that are reffered to as sialyltransferases. It has been found that at least twelve different sialyltransferases are required for preparations of all types of the sialyloligosaccha ride structures so far known (Broquet, P. et al., Int. J. Biochem., 23, 385-389, 1991; and Weinstein, J. et al., J. Biol. Chem., 262, 17735-17743, 1987). Among then, five sialyltransferases were purified and each of the purified enzymes was found to exhibit high specificities to respective acceptor substrates (Sadler, J. et al., J. Bio. Chem., 254, 4434-4443, 1979; Weinstein, J. et al., J. Biol. Chem., 257, 13835-13844, 1982; Rearick, J. et al., J. Biol. Chem., 254, 4444-4451, 1979; and Joziasse, D. H. et al., J. Biol. Chem., 260, 4941-4951, 1985).
With regard to cDNAs encoding the aforementioned sialyltransferases, cDNAs encoding Gal.beta. 1,4GlcNAc.alpha. 2,6-sialyltransferases (Gal.beta. 4GlcNAc-.alpha. 6ST) were cloned from various tissues such as liver (Weinstein, J. et al., J. Biol. Chem., 262, 17735-17743, 1987; Grundmann U. et al., Nucleic Acids Res. 18, 667, 1990; Bast, B. et al., J. Cell. Biol., 116, 423-435, 1992; and Hamamoto, T. et al., Bioorg. and Medic. Chem., 1, 141-145, 1993). In addition, cDNAs encoding Gal.beta. 1,3GalNAc.alpha. 2,3-sialyltransferases (Gal.beta. 3GalNAc-.alpha. 3ST: Gillespie, W. et al., J. Biol. Chem., 267, 21004-21010, 1992; and Lee, Y. et al., Eur. J. Biochem, 216, 377-385, 1993), and a cDNA encoding Gal.beta. 1,3(4)GlcNAc.alpha. 2,3-sialyltransferase (Gal.beta. 3(4)GlcNAc-.alpha. 3ST: Wen, D. X et al., J. Biol. Chem., 267, 21011-21019, 1992) were also cloned.
Furthermore, cDNAs encoding two different types of GalNAc.alpha. 2,6-sialyltransferases (EC 2.4.99.3; GalNAc-.alpha. 6ST) were cloned by the inventors of the present invention, and their soluble proteins were prepared (Kurosawa, N. et al., J. Biol. Chem., 269, pp.1402-1409, 1994; and Kurosawa, N. et al., J. Biol. Chem., 269, pp.19048-19053, 1994). Some other publications also relate to clonings of cDNAs encoding sialyltransferases (e.g. Sasaki, K. et al., J. Biol. Chem., 268, 22782-22787, 1993; and Lee, Y.-C., J. Biol. Chem., 269, 10028-10033, 1994).
Sia.alpha. 2,8Sia-sequences are widely observed in various gangliosides such as GT1a, GD3, and b- and c-series of gangliosides, and are more specifically found in mammal glycoproteins (Troy, F. A., Glycobiology 2, pp.5-23, 1992). It has been reported that Sia.alpha. 2,8Sia-sequences are associated with only two proteins, i.e. the neutral cell adhesion molecule (N-CAM: Edelman, G. M., Annu. Rev. Biochem. 54, pp.135-169, 1985; Cunningham, B. A. et al., Science, 236, pp.799-806, 1987; and Rutishauser, U. et al., Science, 240, pp.53-57, 1988) and the .alpha. subunit of the voltage-gated sodium channels in rat brain (Zuber, C., J. Biol. Chem., 267, pp.9965-9971, 1992).
Recently, the inventors of the present invention cloned an .alpha. 2,8-sialyltransferase, i.e. GD3-synthase (ST8SiaI: Sasaki, K. et al., J. Biol. Chem., 269, pp.15950-15956, 1994), and reported that a developmentally regulated sialyltransferase (STX, ST8SiaII) have N-glycan .alpha.2,8-sialyltransfer activity and polysialic acid synthesizing activity (Kojima, N. et al., FEBS Lett., 360, pp.1-4, 1995, and FEBS Lett., 373, pp.119-122, 1995). However, only two cDNAs encoding .alpha. 2,8-sialyltransferase have been cloned so far, and the substrate specificities of these cloned .alpha. 2,8-sialyltransferases do not give a full explanation as to how all of the Known Sia.alpha. 2,8-Sia sequences in mammal glycolipids and glycoproteins are synthesized.